The ABC (ATP-Binding Cassette) family includes the best known mediators of resistance to anticancer drugs. In particular, MDR (multidrug resistance) pumps actively extrude many types of drugs from cancer cells, thereby conferring resistance to those agents. Arsenite is an increasingly used anticancer agent, while arsenic toxicity is an emerging problem all over the world. Our preliminary results indicate that, unexpectedly, the expression and function of ABCB6 may be an important cellular mechanism to provide arsenite resistance. Despite the established role of ABC transporters in detoxification, this concept remains challenging. ABCB1-MDR1 provides resistance by keeping the intracellular levels of various anticancer agents below a cell-killing threshold. ABCB6 is, however, expressed in the mitochondrial membrane, raising the question as to how its function may convey resistance to the cells. The overall goal of this proposal is to understand how mitochondrial ABC transporters provide drug resistance. The specific hypothesis to be tested is that ABCB6 plays a role in protecting the cells against arsenite-mediated toxicity. This hypothesis is based on the following observations: First, we found that cells selected for resistance to arsenite express higher levels of ABCB6 than parental lines;Second, the overexpression of ABCB6 conferred broad resistance to heavy metals;Third, a pharmacogenomic approach based on the analysis of the NCI60 cell panel suggested the involvement of ABCB6 in drug resistance. Based on these observations, a comprehensive series of studies is proposed to determine the biochemical mechanism by which ABCB6 confers resistance. In particular, the studies proposed are designed to address the following critical questions: First, what substrate(s) does ABCB6 transport? Second, where is ABCB6 located in the cell? Third, what is the functional form of ABCB6? Relevance Although considerable progress has been made in treating cancer over the past decade with a gradual decline in cancer death rates, there are still over 500,000 deaths from cancer in the U.S. each year. Effective treatment of most metastatic cancers requires the use of toxic chemotherapy. Unfortunately, cancer cells may become resistant against cytotoxic agents, often through the elevated activity of ABC transporters, which mediate the energy-dependent efflux of various drugs from cancer cells. This proposal's aim is to elucidate the mechanism and biochemistry of ABCB6, a candidate multidrug transporter.